This disclosure relates generally to expansion and passaging of cells and/or cell aggregates using a stirred tank bioreactor.
A need for large scale pluripotent stem cell culture is emerging for applications in pluripotent stem cell banking (e.g., for induced pluripotent stem cells), commercial production of cells (e.g., GE's Cytiva™ cardiomyocytes), and cell expansion for clinical trials. Advances in feeder-free pluripotent stem cell culture have enabled large scale cell expansion in flasks, on microcarriers (150 to 250 microns in diameter) or on macrocarriers (˜6 mm in diameter) in bioreactors. The use of suspension culture avoids some of the challenges that occur when culturing pluripotent cells on traditional microcarriers including inefficient seeding and release of cells from carriers, physical separation of microcarriers and cells during harvest, and formation of cell-carrier clumping that can lead to phenotypic changes in the cells. Typically, perfusion is used for suspension cultures in bioreactors.
However one challenge in perfusion/suspension culture is how to retain the cells in the bioreactor. Prior art provides some basic separation techniques-1) filtration, 2) gravity sedimentation, and 3) centrifugation. Filtration methods require some means to keep the filter from clogging over the required weeks of operation. A problem with gravity sedimentation is the varying sedimentation characteristics of different cells, the difficulty in scale-up to industrial systems, and difficulty in maintaining sterility. Similarly, centrifugation is routinely used in open cell culture but has found limited application in fully closed system cell culture due to concerns regarding sterility.
There is a need in the field for techniques which reduce human intervention and cross-contamination during the process of culturing cells, including pluripotent stem cells and/or differentiated human cells.